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		<div class="header-chapter">
			<h1 class="heading-1">Brainstorming</h1>

			<div class="box box-gray" style="padding:30px;">
				<p>Since BIOMOD expects the student participants to focus on thinking and brainstorming over various designing techniques at Bio-Molecular level, we discussed a lot of designing techniques for creating 3D DNA Origami Nano-Structures. We had chosen an ultimate goal of creating a 3D replica of The Taj Mahal as our goal.  But initially we were not sure how to go about with the designing. Hence, we looked into a lot of past research and got a lot of ideas. Simply replicating the previously used techniques was one option we had, and which is exactly what we did in Single-shape DNA Origami. However, we were not satisfied with just that. We read a lot of research papers and attempted to combine the designing techniques to create a complex 3D structure and provide techniques to scale the 3D DNA Origami structure.  Here are some of the techniques that we discussed and the ones we finally decided to go with.</p>
			
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<ol class="methodo">
	<li>
		<h3><span class="decimal">1.</span>3D DNA Origami</h3>
		<h4>Packed on a honeycomb lattice using single m13 scaffold</h4>
<div class="description rte"><p>		This technique has previously been used successfully in creating a DNA Robot like 3D structure. Hence, we were quite sure that we would be easily able to replicate this technique. We created the structure using similar principles and called it Single-shape DNA Origami. The design was successfully created but we were not satisfied with the resolution of the structure. We felt the need to scale up the structure so that we could work on its resolution. Hence we looked into other techniques available for 3D DNA Origami.
<br /> 
<br /> Here are some of the snapshots of the structure we designed using this technique:
</p>
		
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		<li>
			<h3><span class="decimal">2.</span>3D DNA Origami packed on a square lattice </h3>
			<h4>Using single m13 scaffold</h4>
	<div class="description rte"><p>This technique is similar to the first technique we used, i.e. Single shape DNA Origami, so we didn’t want to repeat the same process over again. However, there was a reason that we chose a honeycomb lattice over a square lattice for Single-shape DNA Origami. This is because, as stated by Castro et al in their research (6), the square lattice packing rule allows for creating densely packed objects with rectangular features but may require additional effort to eliminate potentially undesired global twist deformations. The honeycomb lattice packing rule by default create straight albeit more porous structures. Thus, we chose the honeycomb lattice for creating our structure. To keep things simple during designing, we went with the honeycomb lattice.
	<br /> 
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			<li>
				<h3><span class="decimal">3.</span>3D DNA Origami with complex hollow curved surface </h3>
				<h4>Using single m13 scaffold</h4>
		<div class="description rte"><p>This technique was motivated by a recent research by Dongran Han et al, where they present their research on DNA Origami with Complex Curvatures in Three-Dimensional Space. We felt that we could similarly create the structure of the Taj Mahal using the principles listed in the research. However, this technique failed. The reason was that the maximum size of m13 scaffold that is available for 3D DNA Origami is around 7.5 – 8 thousand base-pairs in length. This number was way less than what we needed to successfully accomplish our design using this technique, which was around 35000 bps, so we had to abandon this technique too.
		<br /> 
		</p></div> 
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				<li>
					<h3><span class="decimal">4.</span>3D DNA Origami with complex hollow curved surface </h3>
					<h4>Using single m13 scaffold for individual structures and then scaling the structure using staplers </h4>
			<div class="description rte"><p>It was proposed that first we would create individual 3D structures similar to those developed by Hao Yan et al in their research (3), using similar design principles. However, the proposed 3D structures would be a bit different in shape towards the base. Concentric rings would be provided around the base region by extending the same scaffold. 
			<br /> 
			<br /> The following figure puts forward our thoughts-
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			<br /> Once all these structures with a flattened extension of base are created, then a stapling technique that we proposed would come into play. We would need to apply stapler crossovers across two different scaffolds. This would be the glueing mechanism that we could use to join together two three-dimensional structures. Since all the flattened bases of all our 3D structures would lie in the same plane, we could effectively use this multiple-strand stapling technique to scale up the structures. 
			<br /> 
			<br /> Here is a demonstration of the multiple-strand stapling using two simple 2D DNA Origami objects in the same plane.  There would be two types of staplers in the whole structure – those that span over multiple segments of the same strand, and those that connect two strands. The following figure depicts our idea. 
			<br /> As it is shown in the figure, the black staplers create crossovers between two different scaffolds, and this can act as a glue to join two different strands. Similarly, we could theoretically think of joining the 3D structures with a flattened base are that we propose in a similar fashion, since we plan to keep the flattened bases of all the five structures co-planer. However, this technique does not have experimental verification yet.
		</p>
		
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					<li>
						<h3><span class="decimal">5.</span>Multilayer 3D DNA Origami using space-filled structures </h3>
						<h4>Packed on square/honeycomb lattice</h4>
				<div class="description rte"><p>We also attempted to create the Taj Mahal using different Multilayer DNA Origami space-filled structures, and looked into available techniques to join these structures together.  First of all, we created all the individual structures needed for the Taj Mahal (four pillars and central dome), and after this a master plan of the structure was created for future reference, as depicted in the figure. 
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				<br /> Firstly we discussed over how the base could be created. The base could be either a single structure or it can be multiple structures holding the 3D pieces and combining the bases using 3D tile staplers. The following images illustrate both these concepts –
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					<a href="imgs/3d/brainstorming/brainstorm1.png" title="img" target="_blank">
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						<a href="imgs/3d/brainstorming/brainstorm2.png" title="img" target="_blank">
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				<br /> Next we discussed over how to join the five individual structures to the base.
				There were two options for joining the individual structures to the base too -
				
				<br /> 
				<br /> <strong>1. Using slotted cross technique </strong>
				<br /> Slotted cross is a standard existing technique to join two space-filled 3D multilayer origami structures. However, it involves scaffold crossovers, which means that while assigning a sequence, we treat the whole joined structure as one single scaffold. Thus this was not much useful was us since we were looking to scale the structure to more than one scaffold.
				
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					<a href="imgs/3d/brainstorming/cross-slot.png" title="img" target="_blank">
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				<br /> <strong>2. Using lock and key mechanism to fit individual structures onto the base</strong>
				<br /> This was another proposal that was brought up, but the discussion never quite yielded any substantial ideas. Here is a concept illustrated though, a two-faced tetrahedron with one of the sides intended to act as a key which would fit into a corresponding lock and fit in.
			</p>
			
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			<li>
							<h3><span class="decimal">6.</span>Using 3D DNA Bricks</h3>
							<h4>Packed on a honeycomb lattice using single m13 scaffold</h4>
					<div class="description rte"><p>We got the motivation for this idea from a very old research we came across, done at University of Southern California, which was presented in the form of a research paper[22] titled Building Blocks for DNA Self-Assembly. We replicated the 3D DNA Bricks which were proposed in that paper, and have illustrated it in the figure below, but we felt that this technique also wont work because it relied on combining scaffold but didn’t provide much details about how the joining actually happened. It had not been experimentally verified either.
					<br /> 
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